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Apoptosis Antibody Sampler Kit

RMB: 5300

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Catalog# K2001

Apoptosis Antibody Sampler Kit

Application

  • WB

Reactivity

  • Human

General Overview

Kit Components

Product Includes Specification Application Reactivity Mw
Pro Caspase-3[ET1602-39] 20µl WB,IF-Cell,IF-Tissue,IHC-P,IP,FC Human,Monkey Predicted band size: 32 kDa
Cleaved Caspase-3[ET1602-47] 20µl WB,IF-Cell,IHC-P Human Predicted band size: 17 kDa
PARP1[ET1608-56] 20µl WB,IF-Cell,IF-Tissue,IHC-P,FC Human,Mouse,Rat Predicted band size: 113 kDa
Cleaved PARP[ET1608-10] 20µl WB,IF-Cell,IP Human Predicted band size: 89 kDa
Active+Pro Caspase-9[ET1610-95] 20µl WB,IP Human,Mouse Predicted band size: 46 kDa
pro Caspase-7[ET1612-28] 20µl WB,IF-Cell,IF-Tissue,IHC-P Human 34 kDa
Caspase-7[HA723558] 20µl WB,IHC-P Human,Mouse,Rat Predicted band size: 34 kDa
Alpaca anti-Rabbit IgG Fc, Recombinant VHH[HA1031] 100µl IP,ELISA,IHC-P,WB Rabbit

Product Description

The Apoptosis Antibody Sampler Kit designed to provide you with a variety of trial-size antibodies in a convenient and cost-effective format. The kit can be used to detect the full length and cleaved products of Caspase-3, Caspase-7, Caspase-9, and PARP. And also includes secondary reagent for detection of these antibodies.

Product Features

Storage Buffer

1*TBS (pH7.4), 0.05% BSA, 40% Glycerol. Preservative: 0.05% Sodium Azide.

Storage Instructions

Store at +4℃ after thawing. Aliquot store at -20℃. Avoid repeated freeze / thaw cycles.

Background

Apoptosis is a regulated physiological process leading to cell death. Caspases, a family of cysteine acid proteases, are central regulators of apoptosis. Initiator caspases are closely coupled to proapoptotic signals. </br>Once activated, these caspases cleave and activate downstream effector caspases, which in turn cleave cytoskeletal and nuclear proteins like PARP, α-fodrin, DFF, and lamin A and induce apoptosis. Cytochrome c released from mitochondria is coupled to the activation of caspase-9, a key initiator caspase. Proapoptotic stimuli include FasL, TNF-α, DNA damage and ER stress. Fas and TNFR activate caspase-8 and -10, DNA damage leads to the activation of caspase-9 and ER stress leads to the calcium-mediated activation of caspase-12 (3). The inhibitor of apoptosis protein (IAP) family includes XIAP and survivin and functions by binding and inhibiting several caspases. Smac/Diablo, a mitochondrial protein, is released into the cytosol upon mitochondrial stress and competes with caspases for binding of IAPs. The interaction of Smac/Diablo with IAPs relieves the inhibitory effects of IAPs on caspases .

Data Links

Background References

1. Baker, S.J. and Reddy, E.P. (1998) Oncogene 17, 3261-3270.

2. Budihardjo, I. et al. (1999) Annu. Rev. Cell Dev. Biol. 15, 269-290.

3. Nakagawa, T. et al. (2000) Nature 403, 98-103.

4. Deveraux, Q. L. et al. (1998) EMBO J. 17, 2215-2223.

5. Li, F. et al. (1998) Nature 396, 580-584.

6. Du, C. et al. (2000) Cell 102, 33-42.

Images

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Western blot analysis of Caspase-3 on different lysates with Rabbit anti-Caspase-3 antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: HeLa treated with 1μM staurosporine for 3 hours cell lysate<br />Lane 3: Jurkat cell lysate<br />Lane 4: Jurkat treated with 25μM Etoposide for 5 hours cell lysate<br />Lane 5: MCF7 cell lysate (negative)<br />Lane 6: HEK-293 cell lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 32 kDa<br />Observed band size: 32 kDa<br /><br />Exposure time: 3 minutes 20 seconds;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1602-39" style="font-weight: bold;text-decoration: underline;">ET1602-39</a>) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    ☑ Cell treatment (CT)

    Western blot analysis of Caspase-3 on different lysates with Rabbit anti-Caspase-3 antibody (ET1602-39) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: HeLa treated with 1μM staurosporine for 3 hours cell lysate
    Lane 3: Jurkat cell lysate
    Lane 4: Jurkat treated with 25μM Etoposide for 5 hours cell lysate
    Lane 5: MCF7 cell lysate (negative)
    Lane 6: HEK-293 cell lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 32 kDa
    Observed band size: 32 kDa

    Exposure time: 3 minutes 20 seconds;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1602-39) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Western blot analysis of PARP on different lysates with Rabbit anti-PARP antibody (<a href="/products/ET1608-56" style="font-weight: bold;text-decoration: underline;">ET1608-56</a>) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution.<br /><br />Lane 1: HeLa cell lysate (15 µg/Lane)<br />Lane 2: Jurkat cell lysate (15 µg/Lane)<br />Lane 3: NIH/3T3 cell lysate (15 µg/Lane)<br />Lane 4: C2C12 cell lysate (15 µg/Lane)<br />Lane 5: C6 cell lysate (15 µg/Lane)<br />Lane 6: PC-12 cell lysate (15 µg/Lane)<br /><br />Predicted band size: 113 kDa<br />Observed band size: 113 kDa<br /><br />Exposure time: 6 seconds;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1608-56" style="font-weight: bold;text-decoration: underline;">ET1608-56</a>) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1:50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of PARP on different lysates with Rabbit anti-PARP antibody (ET1608-56) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution.

    Lane 1: HeLa cell lysate (15 µg/Lane)
    Lane 2: Jurkat cell lysate (15 µg/Lane)
    Lane 3: NIH/3T3 cell lysate (15 µg/Lane)
    Lane 4: C2C12 cell lysate (15 µg/Lane)
    Lane 5: C6 cell lysate (15 µg/Lane)
    Lane 6: PC-12 cell lysate (15 µg/Lane)

    Predicted band size: 113 kDa
    Observed band size: 113 kDa

    Exposure time: 6 seconds;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1608-56) at 1/2,000 dilution and competitor's antibody at 1/1,000 dilution were used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:50,000 dilution was used for 1 hour at room temperature.

  • <span style="font-weight: bold;">☑ Cell treatment (CT)</span><br /><br />Western blot analysis of Cleaved PARP on different lysates with Rabbit anti-Cleaved PARP antibody (<a href="/products/ET1608-10" style="font-weight: bold;text-decoration: underline;">ET1608-10</a>) at 1/2,000 dilution.<br /><br />Lane 1: HeLa whole cell lysate<br />Lane 2: HeLa treated with 1μM staurosporine for 3 hours whole cell lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 89 kDa<br />Observed band size: 89 kDa<br /><br />Exposure time: 1 minute 9 seconds;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1608-10" style="font-weight: bold;text-decoration: underline;">ET1608-10</a>) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1:50,000 dilution was used for 1 hour at room temperature.

    ☑ Cell treatment (CT)

    Western blot analysis of Cleaved PARP on different lysates with Rabbit anti-Cleaved PARP antibody (ET1608-10) at 1/2,000 dilution.

    Lane 1: HeLa whole cell lysate
    Lane 2: HeLa treated with 1μM staurosporine for 3 hours whole cell lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 89 kDa
    Observed band size: 89 kDa

    Exposure time: 1 minute 9 seconds;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (ET1608-10) at 1/2,000 dilution was used in 5% NFDM/TBST at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1:50,000 dilution was used for 1 hour at room temperature.

  • Western blot analysis of Caspase-9 on different lysates with Rabbit anti-Caspase-9 antibody (<a href="/products/ET1610-95" style="font-weight: bold;text-decoration: underline;">ET1610-95</a>) at 1/1,000 dilution.<br /><br />Lane 1: Hela-si NT cell lysate<br />Lane 2: Hela-si Caspase-9 cell lysate<br /><br />Lysates/proteins at 10 µg/Lane.<br /><br />Predicted band size: 46 kDa<br />Observed band size: 46 kDa<br /><br />Exposure time: 2 minutes;<br /><br />4-20% SDS-PAGE gel.<br /><br /><a href="/products/ET1610-95" style="font-weight: bold;text-decoration: underline;">ET1610-95</a> was shown to specifically react with Caspase-9 in Hela-si NT cells. Weakened band was observed when Hela-si Caspase-9 sample was tested. Hela-si NT and Hela-si Caspase-9 samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (<a href="/products/ET1610-95" style="font-weight: bold;text-decoration: underline;">ET1610-95</a>, 1/1,000) and Loading control antibody (Rabbit anti-GAPDH, <a href="/products/ET1601-4" style="font-weight: bold;text-decoration: underline;">ET1601-4</a>, 1/10,000) were used in 5% BSA at room temperature for 2 hours. Goat Anti-rabbit IgG-HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1:100,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of Caspase-9 on different lysates with Rabbit anti-Caspase-9 antibody (ET1610-95) at 1/1,000 dilution.

    Lane 1: Hela-si NT cell lysate
    Lane 2: Hela-si Caspase-9 cell lysate

    Lysates/proteins at 10 µg/Lane.

    Predicted band size: 46 kDa
    Observed band size: 46 kDa

    Exposure time: 2 minutes;

    4-20% SDS-PAGE gel.

    ET1610-95 was shown to specifically react with Caspase-9 in Hela-si NT cells. Weakened band was observed when Hela-si Caspase-9 sample was tested. Hela-si NT and Hela-si Caspase-9 samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (ET1610-95, 1/1,000) and Loading control antibody (Rabbit anti-GAPDH, ET1601-4, 1/10,000) were used in 5% BSA at room temperature for 2 hours. Goat Anti-rabbit IgG-HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.

  • Western blot analysis of Active+Pro Caspase-9 on different lysates with Rabbit anti-Active+Pro Caspase-9 antibody (<a href="/products/R1308-12" style="font-weight: bold;text-decoration: underline;">R1308-12</a>) at 1/1,000 dilution.<br /><br />Lane 1: Hela-si NT cell lysate<br />Lane 2: Hela-si Caspase-9 cell lysate<br /><br />Lysates/proteins at 10 µg/Lane.<br /><br />Predicted band size: 46 kDa<br />Observed band size: 46 kDa<br /><br />Exposure time: 5 minutes;<br /><br />4-20% SDS-PAGE gel.<br /><br /><a href="/products/R1308-12" style="font-weight: bold;text-decoration: underline;">R1308-12</a> was shown to specifically react with Active+Pro Caspase-9 in Hela-si NT cells. Weakened band was observed when Hela-si Active+Pro Caspase-9 sample was tested. Hela-si NT and Hela-si Active+Pro Caspase-9 samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (<a href="/products/R1308-12" style="font-weight: bold;text-decoration: underline;">R1308-12</a>, 1/1,000) and Loading control antibody (Rabbit anti-GAPDH, <a href="/products/ET1601-4" style="font-weight: bold;text-decoration: underline;">ET1601-4</a>, 1/10,000) were used in 5% BSA at room temperature for 2 hours. Goat Anti-rabbit IgG-HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1:100,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of Active+Pro Caspase-9 on different lysates with Rabbit anti-Active+Pro Caspase-9 antibody (R1308-12) at 1/1,000 dilution.

    Lane 1: Hela-si NT cell lysate
    Lane 2: Hela-si Caspase-9 cell lysate

    Lysates/proteins at 10 µg/Lane.

    Predicted band size: 46 kDa
    Observed band size: 46 kDa

    Exposure time: 5 minutes;

    4-20% SDS-PAGE gel.

    R1308-12 was shown to specifically react with Active+Pro Caspase-9 in Hela-si NT cells. Weakened band was observed when Hela-si Active+Pro Caspase-9 sample was tested. Hela-si NT and Hela-si Active+Pro Caspase-9 samples were subjected to SDS-PAGE. Proteins were transferred to a PVDF membrane and blocked with 5% NFDM in TBST for 1 hour at room temperature. The primary antibody (R1308-12, 1/1,000) and Loading control antibody (Rabbit anti-GAPDH, ET1601-4, 1/10,000) were used in 5% BSA at room temperature for 2 hours. Goat Anti-rabbit IgG-HRP Secondary Antibody (HA1001) at 1:100,000 dilution was used for 1 hour at room temperature.

  • Western blot analysis of Caspase-7 on different lysates with Rabbit anti-Caspase-7 antibody (<a href="/products/HA723558" style="font-weight: bold;text-decoration: underline;">HA723558</a>) at 1/5,000 dilution.<br /><br />Lane 1: HeLa cell lysate<br />Lane 2: HeLa treated with 1μM staurosporine for 3 hours cell lysate<br />Lane 3: C2C12 cell lysate<br />Lane 4: C2C12 treated with 1μM staurosporine for 3 hours cell lysate<br /><br />Lysates/proteins at 20 µg/Lane.<br /><br />Predicted band size: 34 kDa<br />Observed band size: 34/32/20/18 kDa<br /><br />Exposure time: 2 minute 30 seconds; ECL: K1801;<br /><br />4-20% SDS-PAGE gel.<br /><br />Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (<a href="/products/HA723558" style="font-weight: bold;text-decoration: underline;">HA723558</a>) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (<a href="/products/HA1001" style="font-weight: bold;text-decoration: underline;">HA1001</a>) at 1/50,000 dilution was used for 1 hour at room temperature.

    Western blot analysis of Caspase-7 on different lysates with Rabbit anti-Caspase-7 antibody (HA723558) at 1/5,000 dilution.

    Lane 1: HeLa cell lysate
    Lane 2: HeLa treated with 1μM staurosporine for 3 hours cell lysate
    Lane 3: C2C12 cell lysate
    Lane 4: C2C12 treated with 1μM staurosporine for 3 hours cell lysate

    Lysates/proteins at 20 µg/Lane.

    Predicted band size: 34 kDa
    Observed band size: 34/32/20/18 kDa

    Exposure time: 2 minute 30 seconds; ECL: K1801;

    4-20% SDS-PAGE gel.

    Proteins were transferred to a PVDF membrane and blocked with 5% NFDM/TBST for 1 hour at room temperature. The primary antibody (HA723558) at 1/5,000 dilution was used in primary antibody dilution (K1803) at 4℃ overnight. Goat Anti-Rabbit IgG - HRP Secondary Antibody (HA1001) at 1/50,000 dilution was used for 1 hour at room temperature.

  • Immunohistochemical analysis of paraffin-embedded human colon tissue with Rabbit anti-Caspase-7 antibody (<a href="/products/HA723558" style="font-weight: bold;text-decoration: underline;">HA723558</a>) at 1/200 dilution.<br /><br />The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH<sub>2</sub>O and PBS, and then probed with the primary antibody (<a href="/products/HA723558" style="font-weight: bold;text-decoration: underline;">HA723558</a>) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

    Immunohistochemical analysis of paraffin-embedded human colon tissue with Rabbit anti-Caspase-7 antibody (HA723558) at 1/200 dilution.

    The section was pre-treated using heat mediated antigen retrieval with Tris-EDTA buffer (pH 9.0) for 20 minutes. The tissues were blocked in 1% BSA for 20 minutes at room temperature, washed with ddH2O and PBS, and then probed with the primary antibody (HA723558) at 1/200 dilution for 1 hour at room temperature. The detection was performed using an HRP conjugated compact polymer system. DAB was used as the chromogen. Tissues were counterstained with hematoxylin and mounted with DPX.

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